Cell Division and Cell Death

Cell Division and Cell Death

Introduction

• Professor Linda Erskine, a developmental biologist, will be covering cell division and cell death.
• Professor Ferguson will be covering egg to organism and sexual reproduction

Topics Covered

• Cell cycle and its regulation by CDKs and cyclins.
• Impacts of cell cycle dysregulation.
• Mitosis.
• Mechanisms of cell death: necrosis and programmed cell death (apoptosis).

Cellular Environment and Signals

• Cells are constantly bombarded with information, including signaling molecules (e.g., growth factors), physical environment (oxygen levels, matrix stiffness), and nutrient availability.
• Cells integrate these signals to decide whether to:
  • Remain as they are.
  • Undergo cell division (proliferation).
  • Die.
  • Differentiate into a specific cell type.
• Intrinsic properties of the cell also matter, such as specific receptors for growth factors.

Sonic Hedgehog Example:

• A molecule called sonic hedgehog influences the development of our brain and our limb.
• Intrinsic differences in cells allow our body to use the molecule in different ways.
  • In the limb, it determines whether you make a thumb or pinky.
  • In the brain, it triggers the creation of neurons.

Cell Proliferation

• Initiated by extrinsic or intrinsic signals.
• Involves three basic steps:
  • DNA replication.
  • Separation of DNA into two cells.
  • Cytokinesis (splitting the original cell into two new cells).
• Essential for:
  • Reproduction (e.g., budding yeast).
  • Development (e.g., root tissue in plants).
  • Tissue regeneration and repair (e.g., lizard regrowing its tail).
  • Homeostasis (e.g., replacing lost skin cells).

Regulation of Cell Division

• Cell division is tightly regulated, with different cells having different rates of cell division.
  • Early embryonic cells divide rapidly.
  • Differentiated cells (e.g., neurons) do not divide.
• Regulation occurs in the cell cycle.

The Cell Cycle

• The cell cycle consists of:
  • Mitosis (M phase): separation of genetic information and cytokinesis.
  • Interphase (G1, S, and G2 phases).
• The duration of the cell cycle determines how rapidly cells divide.
  • Embryonic cells: ~30 minutes.
  • Rapidly dividing adult cells: ~24 hours.
  • Some cells: weeks or months.

Phases of the Cell Cycle:

• G1 (GAP1):
  • Cell increases in size.
  • Prepares to accommodate more genetic information.
  • Maintains a single copy of its chromosomes.
  • Most variable in length; determines cell cycle duration.
  • Cell commits to DNA replication and mitosis.
• S Phase:
  • DNA replication occurs.
  • Each chromosome forms two identical sister chromatids.
• G2:
  • Preparation for mitosis.
  • Cell assembles structures needed for genetic information separation.
• G0:
  • Quiescent phase where cells are resting.
  • Cells can remain in G0 for extended periods (e.g., quiescent stem cells) or permanently (e.g., differentiated cells like neurons).
  • Cells in G0 can re-enter G1 upon receiving specific signals.

Duration of Cell Cycle Phases (Example):

• In a 24-hour cell cycle:
  • G1: ~11 hours
  • S phase: ~8 hours
  • G2: ~4 hours
  • M phase: ~1 hour
• Mitotic cells are hard to find in tissue because mitosis is relatively short.

Regulation of the Cell Cycle: Cyclins and CDKs

• 2001 Nobel Prize awarded to Leland Hartwell, Tim Hunt, and Sir Paul Nurse for their discovery of key cell cycle regulators.
• Key regulators: cyclins and cyclin-dependent kinases (CDKs).

Cyclin-Dependent Kinases (CDKs):

• Kinases phosphorylate other proteins, changing their activity.
• Regulate cell cycle progression.
• Present in cells all the time (constitutive proteins).
• Inactive unless bound to cyclins.

Cyclins:

• Present cyclically.
• Bind to CDKs, causing a change in CDK shape and activating it.
• The cyclin-CDK complex then phosphorylates specific proteins, impacting the cell cycle.

Mammalian Cyclins and CDKs:

• Different CDKs and cyclins interact in various combinations to form complexes that act in different parts of the cell cycle.
  • CDK1 + cyclin A: regulates G2 to M transition.
  • CDK1 + cyclin B: regulates M phase.
  • CDK2 + cyclin E: regulates G1 to S phase transition.
  • CDK2 + cyclin A: regulates S phase.
  • CDK4/6 + cyclin D: regulates G1 phase.

G1 to S Phase Transition Example: Retinoblastoma Protein (RB)

• RB protein is a negative regulator of the cell cycle.
• When active, RB blocks the cell from going from G1 into S at the restriction point.
• CDK cyclin complex phosphorylates RB protein.
• When phosphorylated, RB becomes inactive, allowing cells to move from G1 into S phase.

Cyclical Presence of Cyclins:

• Cyclins are transient proteins, produced and degraded at specific times during the cell cycle.
• Different cyclins peak in concentration at different phases:
  • Cyclin D: S phase
  • Cyclin E: G1 to S transition
  • Cyclin A: G2
  • Cyclin B: M phase

Summary of Cell Cycle Regulation

• CDKs regulate cell cycle progression but are constitutively present and inactive unless bound to cyclins.
• Cyclins are transiently made during specific parts of the cell cycle, activating CDKs.
• Different cyclin CDK complexes target different proteins, regulating different parts of the cell cycle.
• The presence or absence of cyclins is critical for cell cycle progression.
• Once the complex of cyclin and CDK is done, the cyclin breaks down to allow the CDK to become free again.

Cell Cycle Checkpoints

• Three checkpoints in interphase and one in mitosis.
• Specific triggers cause pausing at these checkpoints.
  • G1: Triggered by DNA damage or unfavorable conditions (lack of nutrients or growth factor stimulation).
  • S phase: Triggered by incomplete replication or DNA damage.
  • G2: Triggered by DNA damage or unduplicated chromosomes.
  • M phase: Triggered by chromosomes not being attached to a spindle.
• The cell pauses to see if it can rectify the problem.
• If the damage can be repaired, the checkpoint is released, and the cell cycle continues.
• If the damage cannot be repaired, the cell undergoes apoptosis.

Regulation of G1 to S Checkpoint Example: p21 Protein

• DNA damage during G1 (e.g., due to radiation) triggers production of p21 protein.
• p21 binds to CDK, preventing cyclin binding and keeping CDK inactive.
• The cell cycle is paused until the p21 is removed.
• If the cell repairs its DNA, p21 breaks down, releasing CDK to interact with cyclin.
• If the DNA cannot be repaired, the cell undergoes apoptosis.

Dysregulation of the Cell Cycle and Cancer

• Consequences of dysregulated cell cycle: cancers.
• Cancer cells ignore checkpoints and lose response to cell cycle controls, dividing continuously.
• Loss of cell cycle control can occur in two ways:
  • Oncogenes:
    • Positive regulators of cell division become overactive, driving unregulated cell division.
    • Examples: growth factors, receptors for growth factors.
    • Mutation in human epidermal growth factor receptor (HER2) in breast cancer.
  • Tumor Suppressors:
    • Negative regulators of cell division become inactive.
    • Example: RB protein.
• Multiple changes are needed for dysregulation to occur.
• Many cancer treatments target the cell cycle by blocking the function of the mitotic spindle, inhibiting growth at the restriction point, blocking DNA replication, or damaging DNA (radiation).
• Problem: these treatments also affect non-cancerous cells, causing side effects (e.g., hair loss).

Mitosis

• Process of segregating DNA and producing two identical cells.
• Continuous process divided into five stages:
  • Prophase
  • Prometaphase
  • Metaphase
  • Anaphase
  • Telophase
• Followed by cytokinesis, which splits the cytoplasm to form two separate cells.

DNA Packaging

• DNA is extensively packaged to fit within the nucleus.
  • DNA double helix wraps around histones to form nucleosomes.
  • Nucleosomes coil together to form chromatin fibers.
  • Chromatin fibers attach to a protein scaffold and loop.
  • Chromatin condenses to form chromatin, then compacted chromosomes.
  • The compacted chromosome that DNA is relatively stable and ready to be separated.

Stages of Mitosis:

• Prophase:
  • DNA condenses to form chromosomes, each consisting of identical paired sister chromatids.
  • Centrosomes move to opposite poles of the cell, anchoring the mitotic spindle.
• Prometaphase:
  • The nuclear envelope breaks down.
  • Mitotic spindle formation continues, connecting each centrosome to the chromosomes.
• Metaphase:
  • Centromeres become aligned in the plane of the cell's equator, forming the metaphase plate.
  • Chromosomes align along this equator.
• Anaphase:
  • Chromatids separate and move to opposite poles of the cell.
  • Regulated by a cell cycle checkpoint.
• Telophase:
  • The spindle breaks down.
  • The nuclear envelope reforms around both newly separated sets of chromosomes.
  • Chromatin decondenses.

The Mitotic Spindle

• Composed of microtubules.
  • Centrosomes: Microtubule-organizing centers to which spindle fibers attach.
  • Polar microtubules:
    • Run from one pole to another.
    • Connect the centrosomes and stabilize the structure.
  • Kinetochore microtubules: Attach the kinetochore to the spindle.
  • Kinetochore: Protein complex that forms a centromeric region of chromosomes and attaches sister chromatids to centrosomes on opposite sides of the cell.
• Motor proteins contract the microtubules apart in anaphase.

Cytokinesis (Animal Cells)

• Separates one cell into two separate cells.
• Begins with furrowing of the cell membrane.
• Driven by actin and myosin contraction, squeezing the cell in its middle.
• Ultimately separates the two cells from each other.
• Each daughter cell contains the same genome, but they may not be identical (different signaling molecules, ribosomes, or fates).

Cell Death

Types of Cell Death:

• Necrosis:
  • Death due to damage.
  • Caused by extremes in temperature, toxins, oxygen or nutrient starvation, or burns.
  • Passive form of cell death.
  • Cells swell up, their membrane breaks down, and they lyse their contents into the organism.
  • Creates an inflammatory response, attracting leukocytes and phagocytes, which can have a collateral effect on the surrounding tissue.
  • Generally detrimental.
• Programmed Cell Death (Apoptosis).
  • Planned cell death.
  • Andrew Wiley coined the term apoptosis, meaning falling off, comparing it to the falling off of leaves from a tree during autumn.
  • Active process.
  • Cells undergo characteristic events:
    • Become detached from their neighbors.
    • Get smaller.
    • Chromatin condenses and is digested into fragments.
    • Blebs break off.
    • The cell breaks up into smaller bits (apoptotic bodies).
    • Apoptotic bodies are engulfed by neighboring cells and their contents are reused.
    • No inflammatory response.
  • Important functions:
    • Eliminates unwanted cells.
    • Balances cell number along with proliferation.
    • Gets rid of cells that have undergone pathogenic change (DNA damage or viral infection).
    • Regulates homeostasis after infection (immune cells) or cells that recognize autoantigens.

Examples of Development.

• During metamorphosis, get a tadpole, it will lose its tail as it becomes a frog, and those tail cells will die off by apoptosis.
• Our limbs develop and have webbing between our fingers. In order for us to get proper fingers, that webbing has to be removed, and that's removed by apoptosis.

Regulation of Apoptosis:

• Tightly regulated by extrinsic and intrinsic signals.
• Signals activate signaling pathways, telling the cell to die.
• Key proteins: caspases and BCL2.

Caspases:

• Proteases that cleave specific targets.
• Found in both the cytoplasm and mitochondria.
• Need to be cleaved to be active.
  • Made as a pro-caspase, then cleaved to make active enzymes.
    • Regulated by an initiator caspase
    • Some caspase cleave caspases, some caspases are going to be effector caspases.

Pathways of Caspase Activation:

• Extrinsic pathway:Signals bind to a death receptor, activating pro-caspases to become active initiator caspases, which then cleave executor caspases, triggering cell death.
• Intrinsic pathway: Release of cytochrome c by mitochondria triggers caspase nine to become active, which then cleaves pro-caspases to form executed caspases, triggering cell death.
• BCL2: Acts to prevent an increase in mitochondrial permeability, preventing the release of cytochrome c.

Conclusion

• Eukaryotic cell cycle and its regulation.
• Mitosis.
• Cell death.